背景:MicroRNAs(miRNA)是重要的非编码RNA实体,通过与靶mRNA结合来影响基因表达和功能,导致mRNA降解或抑制其翻译。miRNA广泛参与多种生物过程,如细胞分化,发展,新陈代谢,和凋亡。此外,miRNA与许多疾病相关,包括癌症.然而,传统的检测技术通常存在灵敏度低等缺点,因此,我们需要开发一种快速高效的检测策略来准确检测miRNAs。
结果:我们开发了一种创新的均相电化学发光(ECL)生物传感器。该生物传感器采用CRISPR/Cas12a基因编辑技术来准确高效地检测microRNA(miRNA)。与传统技术相比,这种生物传感器采用独特的均质检测格式,消除了费力的探针固定步骤,大大简化了检测过程。通过使用两种扩增技术-等温扩增和T7RNA聚合酶扩增-生物传感器提高了测定的灵敏度和特异性,在测定中提供优异的检测性能。这使得可以直接从多种生物样品如细胞裂解物和稀释的人血清中评估miRNA。实验结果令人信服地证明了这种生物传感器的非凡性能,包括其1.27aM的极低检测限,高灵敏度,重现性和稳定性。
结论:我们构建的传感器在区分癌细胞系和非癌细胞系中的应用突出了其早期癌症检测和监测的潜力。这种创新的方法代表了miRNA检测领域的重大进步,提供一个用户友好的,成本效益高,和敏感的解决方案,对临床诊断和患者护理具有广泛的影响,尤其是在即时护理环境中。
BACKGROUND: MicroRNAs (miRNAs) are important non-coding RNA entities that affect gene expression and function by binding to target mRNAs, leading to degradation of the mRNAs or inhibiting their translation. MiRNAs are widely involved in a variety of biological processes, such as cell differentiation, development, metabolism, and apoptosis. In addition, miRNAs are associated with many diseases, including cancer. However, conventional detection techniques often suffer from shortcomings such as low sensitivity, so we need to develop a rapid and efficient detection strategy for accurate detection of miRNAs.
RESULTS: We have developed an innovative homogeneous electrochemiluminescence (ECL) biosensor. This biosensor employs CRISPR/Cas12a gene editing technology for accurate and efficient detection of microRNA (miRNA). Compared to conventional technologies, this biosensor employs a unique homogeneous detection format that eliminates laborious probe fixation steps and greatly simplifies the detection process. By using two amplification techniques - isothermal amplification and T7 RNA polymerase amplification - the biosensor improves the sensitivity and specificity of the assay, providing excellent detection performance in the assay. This makes it possible to evaluate miRNA directly from a variety of biological samples such as cell lysates and diluted human serum. Experimental results convincingly demonstrate the extraordinary performance of this biosensor, including its extremely low detection limit of 1.27 aM, high sensitivity, reproducibility and stability.
CONCLUSIONS: The application of our constructed sensor in distinguishing between cancerous and non-cancerous cell lines highlights its potential for early cancer detection and monitoring. This innovative approach represents a major advancement in the field of miRNA detection, providing a user-friendly, cost-effective, and sensitive solution with broad implications for clinical diagnosis and patient care, especially in point-of-care settings.